1. Field of the Invention
The present invention relates to a process for alleviating environmental difficulties associated with the demilitarization of nitrocellulose base gun propellants. More particularly it provides a method for the bioremediation of soils contaminated with such materials.
2. Description of the Prior Art
The nitrate ester of cellulose, also known as nitrocellulose and cellulose nitrate, is the most commonly used energetic ingredient in gun propellant compositions. Nitrocellulose has a limited shelf life because of an auto catalytic decomposition reaction which can occur over long periods of time while in ambient storage. This reaction becomes dangerous when the stabilizer (diphenylamine), present in the propellant composition, is depleted. Each year large quantities of scrap propellant must be disposed of, but it has long been a problem to safely dispose of nitrocellulose based gun propellants. The currently employed disposal method involves an open air burning of these materials. Unfortunately, both the air borne particulates and the pan residues resulting from open air burning are toxic. When these materials are incinerated the resulting toxic pan residues must be treated as a hazardous waste. Even such treated materials produce toxic particulates and probably will not meet future EPA requirements. Current remediation techniques use mobile incinerators to destroy the energetic materials present in contaminated soils. The soil must be dug up to a depth of six feet and the material incinerated. Obviously, this method is very costly and time consuming. For this reason it is desired to develop a microbial method that could result in the mineralization of this material. Early investigations of the biodegradation of nitrocellulose came to the conclusion that nitrocellulose was not directly attacked by microorganisms, but rather degraded by acidic metabolites resulting from their growth.
A number of chemical processes utilizing inorganic sulfides and hydrides have been investigated for the degradation of nitrocellulose. U.S. Pat. No. 4,814,439 teaches a method for nitrocellulose degradation which uses organic sulfhydryl compounds to release the nitrogen from nitrocellulose in the form of inorganic nitrite ions. Nitrocellulose has been shown to be decomposed by acid treatment and yet more readily by alkaline treatment Wendt, T. M. and A. M. Kaplan, "Chemical-Biological treatment process for cellulose nitrate disposal"; J. Water Poll Control Fed. 48:660-668 (1976) reported that a solution resulting from alkaline hydrolysis could efficiently be treated by a combination of anaerobic and aerobic activated sludge process. Hsieh, H. N. and F. J. Tai, "Anaerobic digestion and acid hydrolysis of nitrocellulose", Proceedings on nitrocellulose-- fines, separation and treatment, U.S. Army--Purdue University, West Lafayette, Ind. pp. 110-121 (1993), have shown that 99% of nitrocellulose could be converted into sugars by a single stage acid hydrolysis at 70.degree. C. for 45-60 minutes. Earlier work carried out on microbial degradation of nitrocellulose has produced conflicting results. Bokorny, T., Chemistry and Technology, Pergamon Press, London, England, p. 313 (1965), showed that molds could grow on nitrocellulose suspended in aqueous medium containing mineral salts. Malenkovic, B. and Jacque, M. Chemistry and Technology, Pergamon Press, London, England, p. 313 (1965) suggested that molds were capable of utilizing the dissolved mineral salts but incapable of attacking nitrocellulose. Brodman, B. W. and M. P. Devine, "Microbial attack of nitrocellulose", J. Appl. Polymer Sci. 26: 997-1000 (1981) showed that Aspergillus fumigatus could utilize nitrocellulose containing 11.11% nitrogen. They also indicated that the organism did not utilize the nitrogen directly from nitrocellulose but rather relied on a hydrolysis reaction for a source of nitrogen. Investigations carried out by Kaplan, D. L., et al, "Denitrification of high nitrate loads--Efficiencies of alternate carbon sources" International Biodeterioration 23: 233-248 (1987), have shown that nitrocellulose was not subjected to direct microbial attack and their studies indicated that chemical pre-treatment of nitrocellulose was necessary to generate a modified denitrated polymer that could be attacked by microorganisms. IL'Inskaya, A. N. et al "Growth of microorganisms on cellulose nitro-esters", Biotekhnologiya, 4:495-500 (1988) showed that Aspergillus fumigatus F-316 was capable of forming reducing sugars from nitrocellulose with a nitrogen content of 11.9% and the fungus utilized the nitrate esters group when subjected to nitrogen deficiency. The same authors further reported the possibility of de-esterification of cellulose nitrate esters by using immobilized A. fumigatus and Pseudomonas fluorescens. IL'Inskaya, O. N. et al, "Decomposition of nitrocellulose by a community of microorganisms immobilized on it", Biol Nauki. 6:87-91 (1988), did not observe any nitrocellulose degradation when six immobilized Pseudomonas sps. and anaerobic cellulolytic bacteria were tested. Duran, M., et al. "Anaerobic biotransformation of nitrocellulose", Proceedings on nitrocellulose--fines, separation and treatment, U.S. Army--Purdue University, West Lafayette, Ind. pp. 92-108 (1993), reported nitrocellulose degradation under anaerobic conditions in a two stage feed reactor. Studies carried out by Hsieh, H. N. and F. J. Tai, "Anaerobic digestion and acid hydrolysis of nitrocellulose" in Proceedings on nitrocellulose--fines, separation and treatment, U.S. Army--Purdue University, West Lafayette, Ind. pp. 110-121 (1993), indicated that only small amount of nitrocellulose could be degraded in a conventional anaerobic digestor.
Under aerobic conditions the present invention has accomplished a 35% nitrocellulose degradation in unoptimized cultural conditions. Significant activities of cellulolytic and denitrifying (nitrate and nitrite reductase) enzymes were detected in growing cultures. The present invention, therefore, provides a more environmentally friendly and less costly route to the demilitarization of nitrate esters and the remediation of soils contaminated by these materials and provides a microbial approach to nitrocellulose degradation. It has now been found that nitrate esters can be biodegraded by a process which uses two specific fungi. This biodegradation can be done either with and without the presence of a surfactant. A surfactant provides an advantage in solubilizing the enzymes produced by the microorganisms. The degradation process results in the conversion of the nitrocellulose to innocuous products. This method provides an efficient, cost effective and safe technique for the destruction of nitrate esters. The invention uses enzymes which are known to be very specific and efficient catalysts and capable of reactions that can not be achieved by conventional chemical or physical methods. Further, these reactions are accomplished at room temperature which makes them ideally suited for use with labile energetic materials. The invention uses enzymes produced by two microorganisms Fusarium solani IFO 31093 and Sclerotium rolfsii ATCC 24459 to accomplish the degradation of nitrate esters. While it has been a commonly held belief that nitrocellulose cannot be degraded under aerobic conditions, new and unexpected results are obtained by this invention whereby nitrocellulose can be aerobically degraded.